NUMBL Interacts with TAK1, TRAF6 and NEMO to Negatively Regulate NF-κB Signaling During Osteoclastogenesis

Abstract

NF-κB signaling is essential for osteoclast differentiation and skeletal homeostasis. We have reported recently that NUMB-like (NUMBL) protein modulates osteoclastogenesis by down regulating NF-κB activation. Herein, we decipher the mechanism underlying this phenomenon. We found that whereas NUMBL mRNA expression decreases upon stimulation of wild type (WT) bone marrow macrophages (BMMs) with RANKL, TAK1 deficiency in these cells leads to increased NUMBL and decreased TRAF6 and NEMO expression. These changes were restored upon WT-TAK1 expression, but not with catalytically inactive TAK1-K63W, suggesting that TAK1 enzymatic activity is required for these events. Forced expression of NUMBL inhibits osteoclast differentiation and function as evident by reduction in all hallmarks of osteoclastogenesis. Conversely, NUMBL-null BMMs, show increased osteoclast differentiation and mRNA expression of osteoclast marker genes. Post-translationally, K48-linked poly-ubiquitination of NUMBL is diminished in TAK1-null BMMs compared to elevated K48-poly-ubiquitination in WT cells, indicating increased stability of NUMBL in TAK1-null conditions. Further, our studies show that NUMBL directly interacts with TRAF6 and NEMO, and induces their K48-poly-ubiquitination mediated proteasomal degradation. Collectively, our data suggest that NUMBL and TAK1 are reciprocally regulated and that NUMBL acts as an endogenous regulator of NF-κB signaling and osteoclastogenesis by targeting the TAK1-TRAF6-NEMO axis.

The adaptor protein NumbL is involved in the control of glucolipotoxicity-induced pancreatic beta cell apoptosis

“…determined that NumbL promotes polyubiquitination of TRAF6 and NEMO in osteoclasts [38]. Overall, NumbL depletion seems to be beneficial for the survival of beta cells under diabetogenic conditions.…”

“…TRAF6 activity depends on its polyubiquitination [16], and it is not clear if TRAF6, being an ubiquitin ligase, ubiquitinates itself or if other factors are involved [15,16,38]. Swarnker et al…”

“…As NumbL down regulation inhibits NF-κB signaling under GLT, we evaluated the mechanism through which NumbL is linked to NF-κB. Several groups have reported that NumbL interacts with TRAF6 and modulates NF-κB signaling [37,38]. Thus, we tested the dynamics of 9 interactions between NumbL and TRAF6 under GLT in beta cells.…”

The Adaptor Protein NumbL Is Involved in the Control of Glucolipotoxicity-Induced Pancreatic Beta Cell Apoptosis

“…As NumbL downregulation inhibits NF-κB signaling under GLT, we evaluated the mechanism through which NumbL is linked to NF-κB. Several groups have reported that NumbL interacts with TRAF6 and modulates NF-κB signaling [ 30 , 31 ]. Thus, we tested the dynamics of the interactions between NumbL and TRAF6 under GLT conditions in beta cells.…”

SARS-CoV-2 Exposed Mesenchymal Stromal Cell from Congenital Pulmonary Airway Malformations: Transcriptomic Analysis and the Expression of Immunomodulatory Genes

“…The inflammatory reaction in MSCs-lung is likely due to the interaction of SARS-CoV-2 with TLRs, whose cascade culminates with the activation of NF-κB: upon activation, NF-κB translocates into the nucleus and binds to DNA to promote the transcription of different genes, proinflammatory cytokines among them. NUMBL , encoding for Numb-like protein, is known to be a negative regulator of NF-κB: in an experiment of osteoclastic differentiation, using cells isolated from the bone marrow of RElA-luc reporter mice, it was clear that NUMBL protein decreased RelA levels, a subunit of NF-κB, after RANKL treatment [ 43 ]. So, it is coherent to find NUMBL downregulated in MSCs-lung, where we suppose the activation of NF-κB, and found it upregulated in MSCs-CPAM, where there should not be activation of NF-κB.…”